Differences In Nicotine Metabolism Research Articles

The Role of Nicotine Metabolic Rate on Nicotine Dependence and Rewarding: Nicotine Metabolism in Chinese Male Smokers and Male Mice.

The exact relationship between nicotine metabolism and dependence is not fully understood but is known to be influenced at a molecular level by genetic factors. A sample comprising 274 Chinese adult male smokers was categorized into groups based on their metabolic rates, namely fast, intermediate, and slow metabolizers. We then measured their smoking topography, evaluated their nicotine dependence, and assessed the rewarding effects. Based on these findings, we proposed the hypothesis that the rate of nicotine metabolism could influence the level of dopamine release which in turn had repercussions on the pleasurable and rewarding effects. To test this hypothesis, male mice were selected with different nicotine metabolic rates that closely resembled in the smoker group. We evaluated their nicotine dependence and rewarding effects through conditioned place preference and withdrawal symptom tests, supplemented with dopamine release measurements. In both animal and human, the slow metabolism group (SMG) required less nicotine to maintain a comparable level of dependence than the fast metabolism group (FMG). The SMG could achieve similar rewarding effects to FMG despite consuming less nicotine. Comparable dopamine levels released were therefore critical in setting the nicotine acquisition behavior in this animal model and also for the smokers tested. Our findings suggested that even within the same ethnicity of established smokers (Chinese Han), differences in nicotine metabolism were an important parameter to modulate the degree of nicotine dependence.

Simultaneous Measurement and Distribution Analysis of Urinary Nicotine, Cotinine, Trans-3′-Hydroxycotinine, Nornicotine, Anabasine, and Total Nicotine Equivalents in a Large Korean Population

Measurement of multiple nicotine metabolites and total nicotine equivalents (TNE) might be a more reliable strategy for tobacco exposure verification than measuring single urinary cotinine alone. We simultaneously measured nicotine, cotinine, 3-OH cotinine, nornicotine, and anabasine using 19,874 urine samples collected from the Korean National Health and Nutrition Examination Survey. Of all samples, 18.6% were positive for cotinine, 17.4% for nicotine, 17.3% for nornicotine, 17.6% for 3-OH cotinine, and 13.2% for anabasine. Of the cotinine negative samples, less than 0.3% were positive for all nicotine metabolites, but not for anabasine (5.7%). The agreement of the classification of smoking status by cotinine combined with nicotine metabolites was 0.982–0.994 (Cohen’s kappa). TNE3 (the molar sum of urinary nicotine, cotinine, and 3-OH cotinine) was most strongly correlated with cotinine compared to the other nicotine metabolites; however, anabasine was less strongly correlated with other biomarkers. Among anabasine-positive samples, 30% were negative for nicotine or its metabolites, and 25% were undetectable. Our study shows that the single measurement of urinary cotinine is simple and has a comparable classification of smoking status to differentiate between current smokers and non-smokers relative to the measurement of multiple nicotine metabolites. However, measurement of multiple nicotine metabolites and TNE3 could be useful for monitoring exposure to low-level or secondhand smoke exposure and for determining individual differences in nicotine metabolism. Geometric or cultural factors should be considered for the differentiation of tobacco use from patients with nicotine replacement therapy by anabasine.

Genetic differences in nicotine sensitivity and metabolism in C57BL/6J and NOD/ShiLtJ mouse strains

Genetic background impacts sensitivity to nicotine's rewarding and aversive effects and metabolism, which influences susceptibility to nicotine addiction. This is important because sensitivity to nicotine influences susceptibility to nicotine addiction. Thus, understanding genetic contribution to nicotine sensitivity can aid in identifying risk factors for nicotine addiction. Genetic variability in addiction phenotypes can be modeled in rodent systems, and comparisons of nicotine sensitivity in inbred mice can identify contributing genetic substrates. Our laboratory has identified differences in nicotine sensitivity in male mice from two inbred mouse strains, C57BL/6J and NOD/ShiLtJ. We found that the NOD/ShiLtJ strain experienced greater nicotine-induced locomotor depression and hypothermia than the C57BL/6J strain. To investigate possible differences in nicotine metabolism between strains, subjects were treated with acute nicotine and serum and urine samples were analyzed using LC-MS/MS to quantify nicotine and metabolites. This analysis revealed that NOD/ShiLtJ mice had similar serum nicotine but lower cotinine and 3′-hydroxycotinine levels after nicotine treatment when compared to C57BL/6J mice. Possible genetic factors mediating strain differences were identified by surveying nicotine sensitivity- and metabolism-related genes within the Mouse Phenome Database SNP retrieval tool. Polymorphisms were found in 15 of the 26 examined gene sequences. Liver expression levels of nicotine metabolism-related genes (Cyp2a5, Cyp2a4, and Aox1) were measured using qPCR. NOD/ShiLtJ mice showed lower expression of Cyp2a5 and Cyp2a4 and greater expression of Aox1 in liver tissue. These data demonstrate complex differences in nicotine sensitivity and metabolism driven by genetic differences between C57BL/6J and NOD/ShiLtJ inbred mouse strains.

P13. Dissecting the Association of CYP2A6 With Nicotine Metabolism in African American Smokers

Individual differences in nicotine metabolism by the liver enzyme CYP2A6 predict smoking behaviours as well as cessation and health outcomes. The nicotine metabolite ratio (NMR; 3’hydroxycotinine/cotinine) is a stable biomarker for nicotine clearance, with demonstrated clinical utility in personalizing cessation treatment. We previously showed that common variants in the CYP2A6 region were strongly associated the NMR in African Americans; in this study, we investigated this association in more detail.

Urinary Cotinine and Cotinine + Trans-3'-Hydroxycotinine (TNE-2) Cut-points for Distinguishing Tobacco Use from Nonuse in the United States: PATH Study (2013-2014).

Determine the overall, sex-, and racially/ethnically-appropriate population-level cotinine and total nicotine equivalents (TNE-2, the molar sum of the two major nicotine metabolites) cut-points to distinguish tobacco users from nonusers across multiple definitions of use (e.g., exclusive vs. polytobacco, and daily vs. non-daily). Using Wave 1 (2013-2014) of the U.S. Population Assessment of Tobacco and Health (PATH) Study, we conducted weighted Receiver Operating Characteristic (ROC) analysis to determine the optimal urinary cotinine and TNE-2 cut-points, stratified by sex and race/ethnicity. For past 30-day exclusive cigarette users, the cotinine cut-point that distinguished them from nonusers was 40.5 ng/mL, with considerable variation by sex (male: 22.2 ng/mL; female: 43.1 ng/mL) and between racial/ethnic groups (non-Hispanic other: 5.2 ng/mL; non-Hispanic black: 297.0 ng/mL). A similar, but attenuated, pattern emerged when assessing polytobacco cigarette users (overall cut-point = 39.1 ng/mL, range = 5.5 ng/mL-80.4 ng/mL) and any tobacco users (overall cut-point = 39.1 ng/mL, range = 4.8 ng/mL-40.0 ng/mL). Using TNE-2, which is less impacted by racial differences in nicotine metabolism, produced a comparable pattern of results although reduced the range magnitude. Because of similar frequency of cigarette use among polytobacco users, overall cut-points for exclusive cigarette use were not substantially different from cut-points that included polytobacco cigarette use or any tobacco use. Results revealed important differences in sex and race/ethnicity appropriate cut-points when evaluating tobacco use status and established novel urinary TNE-2 cut-points. These cut-points may be used for biochemical verification of self-reported tobacco use in epidemiologic studies and clinical trials.

Acute effects of a very low nicotine content cigarette on laboratory smoking lapse: Impacts of nicotine metabolism and nicotine dependence.

Reducing cigarette nicotine content to nonaddictive levels facilitates smoking cessation; however, very low nicotine content cigarettes (VLNCs) may not be equally effective across heterogeneous smokers. We evaluated the impact of acute VLNC smoking versus control (sham puffs) on craving, withdrawal and smoking lapse behaviour and whether genetically influenced differences in nicotine metabolism and individual differences in nicotine dependence moderate observed effects. Thirty-three overnight-abstinent smokers (15 slow vs. 17 normal nicotine metabolizers; 17 low vs. 16 high nicotine dependence) smoked a 0.05-mg nicotine VLNC during one session and took sham VLNC puffs during another session, in a counterbalanced order. Craving and withdrawal were assessed before and after smoking and sham puffing. Next, participants completed the McKee Smoking Lapse Task, which measures ability to resist smoking and quantity of ad libitum smoking. VLNC (vs. sham) reduced craving and withdrawal, increased ability to resist smoking and reduced ad libitum smoking. VLNC-induced reduction in craving for positive reinforcement was greater in slow (vs. normal) metabolizers. Nicotine metabolism did not moderate any other VLNC responses. High-dependence (vs. low-dependence) participants engaged in greater ad lib smoking across VLNC and sham conditions. Nicotine dependence did not moderate VLNC responses. VLNC reduced craving, withdrawal and smoking lapse behaviour. Individual differences in nicotine metabolism and dependence had a minimal impact on VLNC responses; however, VLNCs were less effective at reducing craving for positive reinforcement among normal (vs. slow) metabolizers. These findings suggest that desirable VLNC effects may extend across heterogeneous groups of smokers.

Evaluation of nicotine patch adherence measurement using self-report and saliva cotinine among abstainers in a smoking cessation trial

BackgroundAdherence to nicotine patches relates to cessation. This is the first study to examine the validity of self-reported nicotine patch adherence relative to saliva cotinine. MethodsWe used data from 198 clinical trial participants who received 11 weeks of nicotine patches, self-reported patch use, had saliva cotinine 1-week after the start of treatment assessed, and were not smoking when saliva was collected (CO < 6). Self-reported patch adherence was defined as: 3-day (before saliva collection), 7-day (before saliva collection), 3-week use (7 days before, and 14 days after, saliva collection), and 11-week use (7 days before, and 10 weeks after, saliva collection). Analyses, including receiver operating characteristic curves, considered differences in nicotine metabolism. Sensitivity, specificity and positive (PPV) and negative predictive value (NPV) assessed optimal cotinine cut-point for adherence. ResultsSelf-reported 7-day (r = 0.13) and 3-week (r = 0.13) patch use marginally correlated with week 1 cotinine (p’s = 0.08) but not 3-day or 11-week. Significant area under the curve (AUC) values of 0.67 (95 %CI: 0.55−0.79) and 0.72 (95 %CI: 0.57−0.88) were found using 7-day self-report for the overall sample and for slow metabolizers (p’s<0.01), but not for normal metabolizers. Optimal 1-week cotinine cut-points using 7-day self-report were 170 ng/mL (overall) and 184 ng/mL (slow), with sensitivity = 0.56−0.62, specificity = 0.69−0.78, PPV = 0.96−0.97, and NPV = 0.13−0.14. ConclusionsAmong CO-confirmed abstainers, self-reported patch use and saliva cotinine assessed 1-week into treatment, were modestly correlated and optimal cotinine cut-point differed by rate of nicotine metabolism. Seven-day patch use may be a more valid self-report measure of patch adherence based on cotinine than 3-day, 3-week, or 11-week. Rate of nicotine metabolism may affect this relationship.

The Use of the Nicotine Metabolite Ratio as a Biomarker to Personalize Smoking Cessation Treatment: Current Evidence and Future Directions.

The nicotine metabolite ratio (NMR), a genetically informed biomarker of rate of nicotine metabolism, has been validated as a tool to select the optimal treatment for individual smokers, thereby improving treatment outcomes. This review summarizes the evidence supporting the development of the NMR as a biomarker of individual differences in nicotine metabolism, the relationship between the NMR and smoking behavior, the clinical utility of using the NMR to personalize treatments for smoking cessation, and the potential mechanisms that underlie the relationship between NMR and smoking cessation. We conclude with a call for additional research necessary to determine the ultimate benefits of using the NMR to personalize treatments for smoking cessation. These future directions include measurement and other methodologic considerations, disseminating this approach to at-risk subpopulations, expanding the NMR to evaluate its efficacy in predicting treatment responses to e-cigarettes and other noncigarette forms of nicotine, and implementation science including cost-effectiveness analyses.See all articles in this Special Collection Honoring Paul F. Engstrom, MD, Champion of Cancer Prevention.

Associations of CYP2A6 Gene Polymorphism with Smoking Status Among Jordanians: Gender-Related Differences.

Cytochrome P450 2A6 enzyme (CYP2A6), an essential hepatic enzyme involved in the metabolism of drugs, is responsible for a major metabolic pathway of nicotine. Variation in the activity of polymorphic CYP2A6 alleles has been implicated in inter-individual differences in nicotine metabolism. The objective of the current study was to assess the association between the smoking status and the cytochrome P450 2A6 enzyme (CYP2A6) genotype in Jordanians. In the current study, 218 (117 Male and 101 female) healthy unrelated Jordanian volunteers were recruited. CYP2A6*1B, CYP2A6*4 and CYP2A6*9 were determined and correlated with subject smoking status. *1A/*1A was the most common genetic polymorphism in the overall study population, with no significant frequency differences between smokers and non-smokers. When the population was divided according to gender, only male smokers showed a significant correlation between genotype and smoking status. Considering the CYP2A6*9 genotype, the results showed differences in distribution between smokers and non-smokers, but only women showed a significant association between CYP2A6*9 allele genotype and smoking status. The results of this study show that there is a significant association between CYP2A6*9 genotype and smoking status. They also show that CYP2A6 genotype is significantly influenced by gender.

Genetic determinants of CYP2A6 activity across racial/ethnic groups with different risks of lung cancer and effect on their smoking intensity.

Genetic variation in cytochrome P450 2A6 (CYP2A6) gene is the primary contributor to the intraindividual and interindividual differences in nicotine metabolism and has been found to influence smoking intensity. However, no study has evaluated the relationship between CYP2A6 genetic variants and the CYP2A6 activity ratio (total 3-hydroxycotinine/cotinine) and their influence on smoking intensity [total nicotine equivalents (TNE)], across five racial/ethnic groups found to have disparate rates of lung cancer. This study genotyped 10 known functional CYP2A6 genetic or copy number variants in 2115 current smokers from the multiethnic cohort study [African Americans (AA) = 350, Native Hawaiians (NH) = 288, Whites = 413, Latinos (LA) = 437 and Japanese Americans (JA) = 627] to conduct such an investigation. Here, we found that LA had the highest CYP2A6 activity followed by Whites, AA, NH and JA, who had the lowest levels. Adjusting for age, sex, race/ethnicity and body mass index, we found that CYP2A6 diplotypes were predictive of TNE levels, particularly in AA and JA (P trend < 0.0001). However, only in JA did the association remain after accounting for cigarettes per day. Also, it is only in this population that the lower activity ratio supports lower TNE levels, carcinogen exposure and thereby lower risk of lung cancer. Despite the association between nicotine metabolism (CYP2A6 activity phenotype and diplotypes) and smoking intensity (TNE), CYP2A6 levels did not correlate with the higher TNE levels found in AA nor the lower TNE levels found in LA, suggesting that other factors may influence smoking dose in these populations. Therefore, further study in these populations is recommended.

Abstract 4152: Differences in nicotine metabolism among five racial/ethnic groups with disparate risks for lung cancer: The multiethnic cohort study

Abstract Introduction: Globally, lung cancer is the most common cancer and the leading cause of cancer-related deaths. Smoking is a well-established risk factor for this disease; however, self-reported dose does not fully explain the differences in lung cancer risk observed across ethnic/racial populations. Compared to whites, for the same lifetime number of cigarettes smoked, African Americans (AA) and Native Hawaiians (NH) have substantially higher rates of lung cancer, whereas Japanese Americans (JA) and Latinos (LA) have lower rates of disease. These differences may be due to variations in nicotine metabolism, thereby influencing smoking behavior. Methods: Using overnight or first morning urine samples collected in the Multiethnic Cohort study (MEC), we examined nicotine metabolism among 2,300 cancer-free smokers of five racial/ethnic groups with different lung cancer risks: AA (n=356), NH (n=330), whites (n=444), LA (n=459) and JA (n=711). Total and free urinary nicotine metabolites: nicotine, cotinine, and trans-3-hydroxycotinine (3-OH) were measured by liquid chromatography tandem mass spectrometry. Smoking dose was quantified by urinary nicotine equivalents (NE)=total cotinine + total nicotine + total 3-OH. The respective glucuronidation ratio of each metabolite was calculated as the total metabolite minus the free metabolite divided by the free metabolite. CYP2A6 enzymatic activity ratio was calculated as: (total cotinine + 3-OH)/ NE. To examine the difference in metabolite levels across racial/ethnic groups, covariate-adjusted geometric means were computed for each group from linear regression models, adjusting for age at urine collection, sex, creatinine, BMI, and CYP2A6 activity ratio (for the NE and glucuronides models), cigarettes per day (CPD) (for the NE model) or NE (for the glucuronides and CYP2A6 activity models). Results: We found that NE and nicotine, cotinine and 3-OH glucuronidation and CYP2A6 activity differed by race/ethnicity (p≤0.002). For NE, AA had the highest levels, followed by whites, LA, NH and JA (geometric means=30.9, 28.5, 28.3, 27.2 and 25.7 nmol/mL, respectively). For CYP2A6 activity, the ratio was highest in AA, followed by LA, whites, NH and JA (geometric means= 0.80, 0.81, 0.77, 0.73 and 0.62, respectively). We found that for each racial/ethnic group NE was greater for smokers in the higher compared to the lower tertile of CYP2A6 activity. This trend was significant for all groups, except NH (p-trend's= 0.02 [AA], 0.30 [NH], &amp;lt;0.0001 [W, LA and JA]; p-interaction&amp;lt;0.005). Conclusion: CYP2A6 enzymatic activity affects smoking dose beyond CPD, which in part may explain the relatively lower risk of lung cancer of Japanese Americans. Further investigation of racial differences in lung cancer risk using nicotine biomarkers is warranted. Citation Format: Sung-Shim L. Park, Christian Caberto, Maarit Tiirikainen, Lynne Wilkens, Daniel Stram, Christopher A. Haiman, Stephen S. Hecht, Sharon Murphy, Loic Le Marchand. Differences in nicotine metabolism among five racial/ethnic groups with disparate risks for lung cancer: The multiethnic cohort study. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4152. doi:10.1158/1538-7445.AM2014-4152

Cholinergic Transmission during Nicotine Withdrawal Is Influenced by Age and Pre-Exposure to Nicotine: Implications for Teenage Smoking

Adolescence is a unique period of development characterized by enhanced tobacco use and long-term vulnerability to neurochemical changes produced by adolescent nicotine exposure. In order to understand the underlying mechanisms that contribute to developmental differences in tobacco use, this study compared changes in cholinergic transmission during nicotine exposure and withdrawal in naïve adult rats compared to (1) adolescent rats and (2) adult rats that were pre-exposed to nicotine during adolescence. The first study compared extracellular levels of acetylcholine (ACh) in the nucleus accumbens (NAc) during nicotine exposure and precipitated withdrawal using microdialysis procedures. Adolescent (postnatal day, PND, 28-42) and adult rats (PND60-74) were prepared with osmotic pumps that delivered nicotine for 14 days (adolescents 4.7 mg/kg/day; adults 3.2 mg/kg/day; expressed as base). Another group of adults was exposed to nicotine during adolescence and then again in adulthood (pre-exposed adults) using similar methods. Control rats received a sham surgery. Following 13 days of nicotine exposure, the rats were implanted with microdialysis probes in the NAc. The following day, dialysis samples were collected during baseline and following systemic administration of the nicotinic receptor antagonist mecamylamine (1.5 and 3.0 mg/kg, i.p.) to precipitate withdrawal. A second study compared various metabolic differences in cholinergic transmission using the same treatment procedures as the first study. Following 14 days of nicotine exposure, the NAc was dissected and acetylcholinesterase (AChE) activity was compared across groups. In order to examine potential group differences in nicotine metabolism, blood plasma levels of cotinine (a nicotine metabolite) were also compared following 14 days of nicotine exposure. The results from the first study revealed that nicotine exposure increased baseline ACh levels to a greater extent in adolescent versus adult rats. During nicotine withdrawal, ACh levels in the NAc were increased in a similar manner in adolescent versus adult rats. However, the increase in ACh that was observed in adult rats experiencing nicotine withdrawal was blunted in pre-exposed adults. These neurochemical effects do not appear to be related to nicotine metabolism, as plasma cotinine levels were similar across all groups. The second study revealed that nicotine exposure increased AChE activity in the NAc to a greater extent in adolescent versus adult rats. There was no difference in AChE activity in pre-exposed versus naïve adult rats. In conclusion, our results suggest that nicotine exposure during adolescence enhances baseline ACh in the NAc. However, the finding that ACh levels were similar during withdrawal in adolescent and adult rats suggests that the enhanced vulnerability to tobacco use during adolescence is not related to age differences in withdrawal-induced increases in cholinergic transmission. Our results also suggest that exposure to nicotine during adolescence suppresses withdrawal-induced increases in cholinergic responses during withdrawal. Taken together, this report illustrates important short- and long-term changes within cholinergic systems that may contribute to the enhanced susceptibility to tobacco use during adolescence.

Differences in nicotine metabolism of two Nicotiana attenuata herbivores render them differentially susceptible to a common native predator.

Background Nicotiana attenuata is attacked by larvae of both specialist (Manduca sexta) and generalist (Spodoptera exigua) lepidopteran herbivores in its native habitat. Nicotine is one of N. attenuata's important defenses. M. sexta is highly nicotine tolerant; whether cytochrome P450 (CYP)-mediated oxidative detoxification and/or rapid excretion is responsible for its exceptional tolerance remains unknown despite five decades of study. Recently, we demonstrated that M. sexta uses its nicotine-induced CYP6B46 to efflux midgut-nicotine into the hemolymph, facilitating nicotine exhalation that deters predatory wolf spiders (Camptocosa parallela). S. exigua's nicotine metabolism is uninvestigated.Methodology/Principal FindingsWe compared the ability of these two herbivores to metabolize, tolerate and co-opt ingested nicotine for defense against the wolf spider. In addition, we analyzed the spider's excretion to gain insights into its nicotine metabolism. Contrary to previous reports, we found that M. sexta larvae neither accumulate the common nicotine oxides (cotinine, cotinine N-oxide and nicotine N-oxide) nor excrete them faster than nicotine. In M. sexta larvae, ingestion of nicotine as well as its oxides increases the accumulation of CYP6B46 transcripts. In contrast, S. exigua accumulates nicotine oxides and exhales less (66%) nicotine than does M. sexta. Spiders prefer nicotine-fed S. exigua over M. sexta, a preference reversed by topical or headspace nicotine supplementation, but not ingested or topically-coated nicotine oxides, suggesting that externalized nicotine but not the nicotine detoxification products deter spider predation. The spiders also do not accumulate nicotine oxides.ConclusionsNicotine oxidation reduces S. exigua's headspace-nicotine and renders it more susceptible to predation by spiders than M. sexta, which exhales unmetabolized nicotine. These results are consistent with the hypothesis that generalist herbivores incur costs of detoxification, which include the ecological costs of greater predation risks, in addition to the previously demonstrated energetic, physiological and metabolic costs.

Developmental effects of acute, chronic, and withdrawal from chronic nicotine on fear conditioning

Pre-adolescence and adolescence are developmental periods associated with increased vulnerability for tobacco addiction, and exposure to tobacco during these periods may lead to long-lasting changes in behavioral and neuronal plasticity. The present study examined the short- and long-term effects of nicotine and nicotine withdrawal on fear conditioning in pre-adolescent, adolescent, and adult mice, and potential underlying substrates that may mediate the developmental effects of nicotine, such as changes in nicotinic acetylcholine receptor (nAChR) binding, CREB expression, and nicotine metabolism. Age-related differences existed in sensitivity to the effects of acute nicotine, chronic nicotine and nicotine withdrawal on contextual fear conditioning (no changes in cued fear conditioning were seen); younger mice were more sensitive to the acute effects and less sensitive to the effects of nicotine withdrawal 24h post treatment cessation. Developmental differences in nAChR binding were associated with the effects of nicotine withdrawal on contextual learning. Developmental differences in nicotine metabolism and CREB expression were also observed, but were not related to the effects of nicotine withdrawal on contextual learning 24h post treatment. Chronic nicotine exposure during pre-adolescence or adolescence, however, produced long-lasting impairments in contextual learning that were observed during adulthood, whereas adult chronic nicotine exposure did not. These developmental effects could be related to changes in CREB. Overall, there is a developmental shift in the effects of nicotine on hippocampus-dependent learning and developmental exposure to nicotine results in adult cognitive deficits; these changes in cognition may play an important role in the development and maintenance of nicotine addiction.

Do Biological Differences Help Explain Tobacco-Related Disparities?

Smoking behaviors vary by age, educational level, economic status, gender, race/ethnicity, and geographic location. Results from national surveys in the United States have consistently shown substantial variation in smoking prevalence as well as cigarette consumption rates by these factors.1 Epidemiologic and behavioral research have identified patterns of higher smoking prevalence rates among men, whites and African-Americans, and persons with less than high school education among nonimmigrants.1 Immigrants from Latin America and Asia tend to have lower smoking prevalence rates, especially among women. In addition, nondaily smoking and smoking five or fewer cigarettes per day is common among nonwhite smokers.2 Tobacco researchers have tried to understand the behavioral and social aspects of why smoking rates and related diseases differ by race/ethnicity. The potential role of racial/ethnic differences in nicotine metabolism and response to cessation drug therapy has not been fully explored.

Sex Differences in Susceptibility to Carcinogens

Lung cancer has reached epidemic proportions in women, and is now the most common cause of cancer death among both men and women in the United States. While smoking rates have declined marginally in women, the rising impact of lung cancer in women may imply that women are at higher risk from carcinogens secondary to underlying factors related to sex. These factors include differences in female physiology such as bronchial responsiveness and airway size, sex-based differences in nicotine metabolism via the cytochrome p450 system driven by hormones, and differences in DNA repair capacity, as well as the evolution of cigarettes. These hypotheses will be explored in depth in this article.

Low-level prenatal exposure to nicotine and infant neurobehavior

Objective To examine the association between prenatal exposure to nicotine from tobacco smoke and infant neurobehavior using tobacco biomarkers and a sensitive and comprehensive measure of infant neurobehavior. Study design Participants were 318 infants (206 White, 95 Black, 17 Other) and their mothers. Prenatal tobacco smoke exposure was measured twice during pregnancy and once at delivery using maternal serum cotinine. Infant neurobehavior was assessed with the NICU Network Neurobehavioral Scale at approximately 5 weeks after birth. Results Prenatal tobacco smoke exposure was significantly associated with infant neurobehavior after controlling for important covariates, but the specific behaviors associated with exposure varied by race. In White infants, higher cotinine was associated with increased arousal ( p = .030) and excitability ( p = .034), and decreased self-regulation ( p = .010). In contrast, among Black infants, higher cotinine was associated with decreased arousal ( p = .001), excitability ( p = .021), and special handling required to complete the assessment ( p = .003), and increased self-regulation ( p = .021) and hypotonicity ( p = .016). In secondary analyses, we found racial differences in the effects of postnatal exposure to second hand smoke and low-level prenatal exposure. Conclusions Low-level prenatal tobacco smoke exposure is associated with infant neurobehavior at 5 weeks of age, but the specific effects differ by race. These effects may reflect racial differences in nicotine metabolism that are similar to differences reported in adult and child studies of tobacco.

Inhibitory Effects of Neurotransmitters and Steroids on Human CYP2A6

Human CYP2A6 catalyzes the metabolism of nicotine, cotinine, and coumarin as well as some pharmaceutical drugs. CYP2A6 is highly expressed in liver and, also, in brain and steroid-related tissues. In this study, we investigated the inhibitory effects of neurotransmitters and steroid hormones on CYP2A6 activity. We found that coumarin 7-hydroxylation and cotinine 3'-hydroxylation by recombinant CYP2A6 expressed in baculovirus-infected insect cells were competitively inhibited by tryptamine (both K(i) = 0.2 microM), serotonin (K(i) = 252 microM and 167 microM), dopamine (K(i) = 49 microM and 22 microM), and histamine (K(i) = 428 microM and 359 microM). Cotinine formation from nicotine was inhibited by tryptamine (K(i) = 0.7 microM, competitive), serotonin (K(i) = 272 microM, noncompetitive), dopamine, noradrenaline, and adrenaline (K(i) = 11 microM, 54 microM, and 81 microM, uncompetitive). Estrogens (K(i) = 0.6-3.8 microM), androgens (K(i) = 60-149 microM), and corticosterone (K(i) = 36 microM) also inhibited cotinine formation, but coumarin 7-hydroxylation and cotinine 3'-hydroxylation did not. Nicotine-Delta(5'(1'))-iminium ion formation from nicotine was not affected by these steroid hormones, indicating that the inhibition of cotinine formation was due to the inhibitory effects on aldehyde oxidase. The nicotine-Delta(5'(1'))-iminium ion formation was competitively inhibited by tryptamine (K(i) = 0.3 microM), serotonin (K(i) = 316 microM), dopamine (K(i) = 66 microM), and histamine (K(i) = 209 microM). Thus, we found that some neurotransmitters inhibit CYP2A6 activity, being related with inter- and intraindividual differences in CYP2A6-dependent metabolism. The inhibitory effects of steroid hormones on aldehyde oxidase may also contribute to interindividual differences in nicotine metabolism.

Characterization and Comparison of Nicotine and Cotinine Metabolism in Vitro and in Vivo in DBA/2 and C57BL/6 Mice

DBA/2 and C57BL/6 are two commonly used mouse strains that differ in response to nicotine. Previous studies have shown that the nicotine-metabolizing enzyme CYP2A5 differs in coumarin metabolism between these two strains, suggesting differences in nicotine metabolism. Nicotine was metabolized to cotinine in vitro by two enzymatic sites. The high-affinity sites exhibited similar parameters (Km, 10.7 +/- 4.8 versus 11.4 +/- 3.6 microM; Vmax, 0.58 +/- 0.18 versus 0.50 +/- 0.07 nmol/min/mg for DBA/2 and C57BL/6, respectively). In vivo, the elimination half-lives of nicotine (1 mg/kg, s.c.) were also similar between DBA/2 and C57BL/6 mice (8.6 +/- 0.4 versus 9.2 +/- 1.6 min, respectively); however, cotinine levels were much higher in DBA/2 mice. The production and identity of the putative cotinine metabolite 3'-hydroxycotinine in mice was confirmed by liquid chromatography/mass spectrometry/mass spectrometry. The in vivo half-life of cotinine (1 mg/kg, s.c.) was significantly longer in the DBA/2 mice compared with the C57BL/6 mice (50.2 +/- 4.7 versus 37.5 +/- 9.6 min, respectively, p < 0.05). The in vitro metabolism of cotinine to 3'-hydroxycotinine was also less efficient in DBA/2 than C57BL/6 mice (Km, 51.0 +/- 15.6 versus 9.5 +/- 2.1 microM, p < 0.05; Vmax, 0.10 +/- 0.01 versus 0.04 +/- 0.01 nmol/min/mg, p < 0.05, respectively). Inhibitory antibody studies demonstrated that the metabolism of both nicotine and cotinine was mediated by CYP2A5. Genetic differences in Cyp2a5 potentially contributed to similar nicotine but different cotinine metabolism, which may confound the interpretation of nicotine pharmacological studies and studies using cotinine as a biomarker.

Characterization of the novel CYP2A6*21 allele using in vivo nicotine kinetics

The impact of CYP2A6*21 (K476R) on in vivo nicotine metabolism and disposition was investigated. A two-step allele-specific PCR assay was developed to detect the 6573A>G single nucleotide polymorphism (SNP) in CYP2A6*21. Nicotine metabolism phenotypes from a previously described intravenous labeled nicotine and cotinine infusion study [1] was used to assess the impact of CYP2A6*21. Genomic DNA samples from 222 (111 monozygotic and dizygotic twin pairs) Caucasian subjects were genotyped for CYP2A6 alleles (CYP2A6*1X2, -*1B, -*2, -*4, -*7, -*9, -*10, -*12, and -*21). The pharmacokinetic parameters were compared between individuals with no detected CYP2A6 variants (CYP2A6*1/*1, n = 163) and individuals heterozygous for the CYP2A6*21 allele (CYP2A6*1/*21, n = 9). The frequency of the CYP2A6*21 allele was found to be 2.3% in Caucasians (n = 5/222 alleles, evaluated in one twin from each twin pair). In vivo pharmacokinetic parameters, such as nicotine clearance (1.32+/-0.37 vs. 1.18+/-0.20 L/min), fractional clearance of nicotine to cotinine (1.02+/-0.36 vs. 0.99+/-0.23 L/min), nicotine half-life (111+/-37 vs. 116+/-29 min), and the trans-3'-hydroxycotinine to cotinine ratio (1.92+/-1.0 vs. 1.55+/-0.58) indicated no substantial differences in nicotine metabolism between those without the variant (CYP2A6*1/*1, n = 163) and those with the variant (CYP2A6*1/*21, n = 9), respectively. CYP2A6*21 does not have a detectable impact on nicotine metabolism in vivo. Our data suggest that CYP2A6*21 may not be important for future studies of nicotine metabolism and the resulting impacts on smoking behaviors.